4-(Phenyl-piperidin-4-ylidene-methyl)-benzamide derivatives and their use for the treatment of pain, anxiety or gastrointestional disorders

ABSTRACT

Compounds of general formula I R 1  is selected from any one of phenyl, pyridinyl, pyrroloyl, thienyl, furanyl, imidazolyl, triazolyl, thiazolyl and pyridine N-oxide; where each R 1  phenyl ring and R 1  heteroaromatic ring may optionally and independently be further substituted by 1, 2 or 3 substituents selected from straight and branched C 1 –C 6  alkyl, NO 2 , CF 3 , C 1 –C 6  alkoxy, chloro, fluoro, bromo, and iodo. The substitutions on the phenyl ring and on the heteroaromatic ring may take place in any position on said ring systems; are disclosed and claimed in the present application, as well as salts and pharmaceutical compositions comprising the novel compounds and their use in therapy, in particular in the management of pain, anxiety and functional gastrointestinal disorders.

FIELD OF THE INVENTION

The present invention is directed to novel compounds, to a process fortheir preparation, their use and pharmaceutical compositions comprisingthe novel compounds. The novel compounds are useful in therapy, and inparticular for the treatment of pain, anxiety and functionalgastrointestinal disorders.

BACKGROUND OF THE INVENTION

The δ receptor has been identified as having a role in many bodilyfunctions such as circulatory and pain systems. Ligands for the δreceptor may therefore find potential use as analgesics, and/or asantihypertensive agents. Ligands for the δ receptor have also been shownto possess immunomodulatory activities.

The identification of at least three different populations of opioidreceptors (μ, δ and κ) is now well established and all three areapparent in both central and peripheral nervous systems of many speciesincluding man. Analgesia has been observed in various animal models whenone or more of these receptors has been activated.

With few exceptions, currently available selective opioid δ ligands arepeptidic in nature and are unsuitable for administration by systemicroutes. One example of a non-peptidic δ-agonist is SNC80 (Bilsky E. J.et al., Journal of Pharmacology and Experimental Therapeutics, 273(1),pp. 359–366 (1995)). There is however still a need for selectiveδ-agonists having not only improved selectivity, but also an improvedside-effect profile.

Thus, the problem underlying the present invention was to find newanalgesics having improved analgesic effects, but also with an improvedside-effect profile over current μ agonists, as well as having improvedsystemic efficacy.

Analgesics that have been identified and are existing in the prior arthave many disadvantages in that they suffer from poor pharmacokineticsand are not analgesic when administered by systemic routes. Also, it hasbeen documented that preferred δ agonist compounds, described within theprior art, show significant convulsive effects when administeredsystemically.

We have now found certain compounds that exhibit surprisingly improvedproperties, i.a. improved δ-agonist potency, in vivo potency,pharmacokinetic, bioavailability, in vitro stability and/or lowertoxicity.

Outline of the Invention

The novel compounds according to the present invention are defined bythe formula I

whereinR¹ is selected from any one of

where each R¹ phenyl ring and R¹ heteroaromatic ring may optionally andindependently be further substituted by 1, 2 or 3 substituentsindependently selected from straight and branched C₁–C₆ alkyl, NO₂, CF₃,C₁–C₆ alkoxy, chloro, fluoro, bromo, and iodo. The substitutions on thephenyl ring and on the heteroaromatic ring may take place in anyposition on said ring systems;

A further embodiment of the present invention is a compound according tofigure I wherein R¹ is as defined above and each R¹ phenyl ring and R¹heteroaromatic ring may independently be further substituted by a methylgroup

A further embodiment of the present invention is a compound according tofigure I wherein R¹ is phenyl, pyrrolyl, pyridinyl, thienyl or furanyl,optionally with 1 or 2 of the preferred substituents on the R¹ phenyl orR¹ heteroaromatic ring.

Another embodiment of the present invention is a compound according tofigure I wherein R¹ is phenyl, pyrrolyl or pyridinyl, optionally with 1or 2 of the preferred substituents on the R¹ phenyl or R¹ heteroaromaticring.

Another embodiment of the present invention is a compound according tofigure I wherein R¹ is thienyl or furanyl, optionally with 1 or 2 of thepreferred substituents on the R¹ heteroaromatic ring.

When the R¹ phenyl ring and the R¹ heteroaromatic ring(s) aresubstituted, the preferred substituents are independently selected fromanyone of CF₃, methyl, iodo, bromo, fluoro and chloro.

Reaction step g in Scheme 1, vide infra, is performed by reacting anintermediate compound of the general formula II

wherein PG is a urethane or urethane protecting group, such as Boc andCBZ or benzyl or substituted benzyl protecting group, such as2,4-dimethoxybenzyl, with 3-aminophenyl boronic acid, using a palladiumcatalyst, e.g. Pd(PPh₃)₄, in the presence of a base, e.g. Na₂CO₃, togive the compounds of general formula III

which is thereafter deprotected, under standard conditions and alkylatedunder reductive conditions with a compound of the general formula R¹—CHOto give compounds of the general formula I.

The novel compounds of the present invention are useful in therapy,especially for the treatment of various pain conditions such as chronicpain, neuropathic pain, acute pain, cancer pain, pain caused byrheumatoid arthritis, migraine, visceral pain etc. This list shouldhowever not be interpreted as exhaustive.

Compounds of the invention are useful as immunomodulators, especiallyfor autoimmune diseases, such as arthritis, for skin grafts, organtransplants and similar surgical needs, for collagen diseases, variousallergies, for use as anti-tumour agents and anti viral agents.

Compounds of the invention are useful in disease states wheredegeneration or dysfunction of opioid receptors is present or implicatedin that paradigm. This may involve the use of isotopically labelledversions of the compounds of the invention in diagnostic techniques andimaging applications such as positron emission tomography (PET).

Compounds of the invention are useful for the treatment of diarrhoea,depression, anxiety and stress-related disorders such as post-traumaticstress disorders, panic disorder, generalized anxiety disorder, socialphobia, and obesessive compulsive disorder; urinary incontinence,various mental illnesses, cough, lung oedema, various gastro-intestinaldisorders, e.g. constipation, functional gastrointestinal disorders suchas Irritable Bowel Syndrome and Functional Dyspepsia, Parkinson'sdisease and other motor disorders, traumatic brain injury, stroke,cardioprotection following miocardial infarction, spinal injury and drugaddiction, including the treatment of alcohol, nicotine, opioid andother drug abuse and for disorders of the sympathetic nervous system forexample hypertension.

Compounds of the invention are useful as an analgesic agent for useduring general anaesthesia and monitored anaesthesia care. Combinationsof agents with different properties are often used to achieve a balanceof effects needed to maintain the anaesthetic state (eg. amnesia,analgesia, muscle relaxation and sedation). Included in this combinationare inhaled anaesthetics, hypnotics, anxiolytics, neuromuscular blockersand opioids.

Also within the scope of the invention is the use of any of thecompounds according to the formula I above, for the manufacture of amedicament for the treatment of any of the conditions discussed above.

A further aspect of the invention is a method for the treatment of asubject suffering from any of the conditions discussed above, whereby aneffective amount of a compound according to the formula I above, isadministered to a patient in need of such treatment.

A further aspect of the present invention is intermediates of thegeneral formula II and III,

wherein PG is a urethane protecting group such as Boc or CBZ, or abenzyl or substituted benzyl protecting group, such as2,4-dimethoxybenzyl.Methods of Preparation

EXAMPLES

The invention will now be described in more detail by the followingSchemes and Examples, which are not to be construed as limiting theinvention.

Example 1N,N-diethyl-4-(3-aminolphenyl-piperidin-4-ylidene-methyl)-benzamide(Compound 7)

(i) Preparation of4-(4-Methoxycarbonyl-benzylidene)-piperidine-1-carboxylic AcidTert-Butyl Ester.

A mixture of starting material 1 (11.2 g, 49 mmol) and trimethylphosphite (25 mL) was refluxed under N₂ for 5 hrs. Excess trimethylphosphite was removed by co-distillation with toluene to give compound 2in quantitative yield:

¹H NMR (CDCl₃) δ 3.20 (d, 2H, J=22 Hz, CH₂), 3.68 (d, 3H 10.8 Hz, OCH₃),3.78 (d, 3H, 11.2 Hz, OCH₃), 3.91 (s, 3H, OCH₃), 7.38 (m, 2H, Ar—H),8.00 (d, 2H, J=8 Hz, Ar—H).

(ii) 4-(4-Methoxycarbonyl-benzylidene)-piperidine-1-carboxylic AcidTert-Butyl Ester (Compound 3)

To a solution of 2 in dry THF (200 mL) was added dropwise lithiumdiisopropylamide (32.7 mL 1.5 M in hexanes, 49 mmol) at −78° C. Thereaction mixture was then allowed to warm to room temperature prior toaddition of N-tert-butoxycarbonyl-4-piperidone (9.76 g, 49 mmol in 100mL dry TEIF). After 12 hrs, the reaction mixture was quenched with water(300 mL) and extracted with ethyl acetate (3×300 mL). The combinedorganic phases were dried over MgSO₄ and evaporated to give a crudeproduct, which was purified by flash to provide 3 as a white solid (5.64g, 35%):

IR (NaCl) 3424, 2974, 2855, 1718, 1688, 1606, 1427, 1362, 1276 cm⁻¹;

¹H NMR (CDCl₃) δ 1.44 (s, 9H), 2.31 (t, J=5.5 Hz, 2H), 2.42 (t, J=5.5Hz, 2H), 3.37 (t, J=5.5 Hz, 2H), 3.48 (t, J=5.5 Hz, 2H), 3.87 (s, 3H,OCH₃), 6.33 (s, 1H, CH), 7.20 (d j=6.7 Hz, 2H, Ar—H), 7.94 (d, J=6.7 Hz,2H, Ar—H); ¹³C NMR (CDCl₃) δ 28.3, 29.2, 36.19, 51.9, 123.7, 127.8,128.7, 129.4, 140.5, 142.1, 154.6, 166.8.

(iii) Preparation of4-Bromo-4-[bromo-(4-methoxycarbonyl-phenyl)-methyl]-piperidine-1-carboxylicAcid Tert-Butyl Ester (Compound 4)

To a mixture of 3 (5.2 g, 16 mmol) and K₂CO₃ (1.0 g) in drydichloromethane (200 mL) was added a solution of bromine (2.9 g, 18mmol) in 30 mL CH₂Cl₂ at 0° C. after 1.5 hrs at room temperature, thesolution after filtration of K₂CO₃ was condensed. The residue was thendissolved in ethyl acetate (200 mL), washed with water (200 mL), 0.5 MHCl (200 mL) and brine (200 mL), and dried over MgSO₄. Removal ofsolvents provided a crude product, which was recrystallized frommethanol to give 4 as a white solid (6.07 g, 78%):

IR (NaCl) 3425, 2969, 1725, 1669, 1426, 1365, 1279, 1243 cm⁻¹;

¹H NMR (CDCl₃) δ 1.28 (s, 9H), 1.75 (m, 1H), 1.90 (m, 1H), 2.1 (m, 211),3.90 (s, 3H, OCH₃), 4.08 (br, 3H), 7.57 (d, J=8.4 Hz, 2H, Ar—H) 7.98 (d,J=8.4 Hz, 2H, Ar—H);

¹³C NMR (CDCl₃) δ 28.3, 36.6, 38.3, 40.3, 52.1, 63.2, 72.9, 129.0,130.3, 130.4, 141.9, 154.4, 166.3.

(iv) 4-[bromo-(4-carboxy-phenyl)-methylene]-piperidine-1-carboxylic AcidTert-Butyl Ester (Compound 5)

A solution of 4 (5.4 g 11 mmol) in methanol (300 mL) and 2.0 M NaOH (100mL) was heated at 40° C. for 3 hrs. The solid was collected byfiltration, and dried overnight under vacuum. The dry salt was dissolvedin 40% acetonitrile/water, and was adjusted to pH 2 using concentratedHCl. Product 5 (3.8 g, 87%) was isolated as a white powder byfiltration:

¹H NMR (9DCl₃) δ 1.45 (s, 9H, ^(t)Bu), 2.22 (dd, J=5.5 Hz, 6.1 Hz, 2H),6.1 Hz, 2H), 3.34 (dd, J=5.5 Hz, 6.1 Hz, 211), 3.54 (dd, J=5.5 Hz, 6.1Hz, 2H), 7.35 (d, J=6.7 Hz, 2H, Ar—H), 8.08 (d, J=6.7 Hz, 2H, Ar—H); ¹³CNMR (CDCl₃) δ 28.3, 31.5, 34.2, 44.0, 115.3, 128.7, 129.4, 130.2, 137.7,145.2, 154.6, 170.3;

(v)4-[bromo-(4-diethylcarbamoyl-phenyl)-methylene]-piperidine-1-carboxylicAcid Tert-Butyl Ester (Compound 6)

To a solution of compound 5 (1.0 g, 2.5 mmol) in dry dichloromethane (10mL) at −20° C. was added isobutylchloroformate (450 mg, 3.3 mmol). After20 min at −20° C. diethylamine (4 mL) was added and the reaction wasallowed to warm to room temperature. After 1.5 hrs the solvents wereevaporated and the residue was partitioned between ethyl acetate andwater. The organic phase was washed with brine and dried over MgSO₄.Removal of solvents provided a crude product, which was purified byflash chromatography to give compound 6 as white needles (800 mg, 73%):IR (NaCl) 3051, 2975, 1694, 1633, 1416, 1281, 1168, 1115 cm⁻¹;

¹H NMR (CDCl₃) δ 1.13 (br, 3H, CH₃), 1.22 (br, 3H, CH₃), 1.44 (s, 9H,_(t)Bu), 2.22 (t, J=5.5 Hz, 2H), 2.62 (t, J=5.5 Hz, 2H), 3.33 (m, 4H),3.55 (m, 4H), 7.31 (d, J=8.0 Hz, 2H, Ar—H), 7.36 (d, J=8.0 Hz, 2H,Ar—H); ¹³C NMR (CDCl₃) δ 12.71, 14.13, 28.3, 31.5, 34.2, 39.1, 43.2,79.7, 115.9, 126.3, 129.3, 136.8, 137.1, 140.6, 154.6, 170.5.

(vi) Preparation ofN,N-diethyl-4-(3-hydroxylphenyl-piperidin-4-ylidene-methyl)-benzamide(Compound 7)

To a flask containing vinyl bromide (6) (8.5 g, 18.9 mM) is added xylene(120 mL), ethanol (80 mL) and 3-aminophenylboronic acid (1.5 eq). Thesolution is degassed for 30 minutes, then aqueous sodium carbonate (2N,29 mL, 3.0 eq, degassed for 30 minutes) is added via cannula. Thenpalladium tetrakistriphenylphosphine (9.075 eq) is added. The reactionmixture is degassed for 10 minutes and heated to 80° C. overnight. Thereaction is cooled, diluted with water and filtered through a pad ofdiatomaceous earth. The organics are removed and the aqueous extractedwith ether (2×50 mL). The combined organic extract is dried withanhydrous magnesium sulfate, filtered and concentrated. The residue wasused crude for the next transformations.

Example 2N,N-diethyl-4-(3-aminophenyl-N-benzyl-piperidin-4-ylidene-methyl)-benzamide(Compound 9)

(i) Preparation ofN,N-diethyl-4-(bromo-N-benzyl-piperidin-4-ylidene-methyl)-benzamide(Compound 8)

Compound 6, prepared in Example 1 (v) above (6.122 g, 13.4 mmoL), wastreated with TFA (13 mL) in dichloromethane (80 mL) at room temperature.After 2 h, the reaction mixture was washed with 2M sodium hydroxide (25mL) and the organic layer was separated. The organic layer was dried(MgSO), filtered and concentrated. The residue was dissolved indichloromethane (120 mL), cooled to 0° C. and benzyl bromide (1.8 mL,15.1 mmol) and triethylamine (5.7 mL, 41.0 mmol) were added. Thereaction was gradually warmed to room temperature and after 20 hours thereaction was washed with water (1×100 mL). The organic layer was dried(MgSO₄), filtered and concentrated. Purification by flashchromatography, eluting 50 to 60% ethyl acetate in hexanes gave 3.80 gof product (64% yield).

¹H NMR (CDCl₃) δ 1.13 (br, 3H, CH₃), 1.23 (br, 3H, CH₃), 2.28 (m, 2H),2.55 (m, 2H), 2.69 (m, 2H), 3.27 (m, 2H), 3.53 (br, 4H), 7.20–7.40 (m,9H, Ar

(ii)N,N-diethyl-4-(3-aminophenyl-N-benzyl-piperidin-4-ylidene-methyl)-benzamide(Compound 9)

To a flask containing 8.5 g of vinyl bromide (8) was added 120 mLxylene, 80 mL ethanol and 3.96 g 3-aminophenyl boronic acid (1.5 eq).The solution was degassed for 30 minutes then 29.0 mL 2N sodiumcarbonate (3.0 eq) (degassed for 30 minutes) was added via cannula. Thenpalladium tetrakistriphenylphosphine (1.67 g, 0.075 eq) was added. Thereaction mixture was degassed for 10 minutes, then warmed to 80° C. for17 hours. The mixture was then cooled, diluted with water and filteredthrough diatomaceous earth. The organics were removed and the aqueousextracted with ether (2×). The combined organics were dried withanhydrous magnesium sulfate, filtered and concentrated. Residue waspurified by flash chromatography eluting with 2% methanol to 4% methanolin dichloromethane. The product (8.14 g, 93%) was obtained as as orangefoam. Residue was dissolved in 80 mL ether and 40 mL HCl/ether wasadded. The suspension was concentrated after 30 minutes and solid wasdried under high vacuum.

Examples 3–9

Additional Examples 3–9 were prepared by following the general syntheticprocedure below.

To a solution of compound 7 in dry tetrahydrofuran (THF) is added thealdehyde (1–1.5 eq), followed by sodium triacetoxyborohydride (1–1.6eq). The reaction is stirred at room temperature under a nitrogenatmosphere for an extended period of time (6–48 hours) to ensurecomplete reaction. The reaction mixture is then subjected to a standardwork-up procedure and standard purification. The amount of THF is notcrucial. An amount corresponding to about 30 mL per gram of amine ispreferred.

The procedure described below for Example 3 is typical.

Example 3

To a solution of amine 7 (540 mg, 1.48 mmol, 1.0 equiv.) intetrahydrofuran (20 ml) at room temperature was added 2-pyridinecarboxaldehyde (170 μl, 1.38 mmol, 1.2 equiv.). After stirring for 10minutes sodium triacetoxyborohydride (410 mg, 1.93 mmol, 1.3 equiv.) wasadded to the solution. After stirring overnight, the reaction mixturewas diluted with dichloromethane (10 ml) and 2M aqueous sodium hydroxidesolution (15 ml). The phases were separated and the aqueous phase isback-extracted with dichloromethane three times (15 ml). The organicphases were combined, dried with sodium sulfate, filtered andconcentrated under reduced pressure. The crude product was purified byreverse phase preparative HPLC (gradient:10% to 50% B in A, A:0.1% TFAin water; B: 0.1% TFA in acetonitrile). The fraction was concentratedunder reduced pressure and neutralized to pH=11 with 2M aqueous sodiumhydroxyde solution. The mixture is then extracted twice with ethylacetate (30 ml). The organic phases are combined, dried with sodiumsulfate, filtered. To this mixture was added 1M HCl solution in diethylether (4 ml, ca. 3.5 equiv.). The resulting mixture was thenconcentrated under reduced pressure. The white solid was triturated withdiethyl ether and concentrated under reduced pressure to yield 135 mg,(19% yield)

Additional examples were prepared analogously. Analytical data for thesynthetic Examples is shown in Table 1 below.

TABLE 1 Analytical data for synthetic Examples. Ex. # R¹ Name NMRdata(400MHZ CD₃OD) 2

4-[1-(3-Amino-phenyl)-1-(1-benzyl-piperidin-4-ylidene)-methyl]-N,N-diethyl-benzamide.7.48–7.56(m, 6H, Ar-H); 7.26–7.38(m, 6H,Ar-H); 7.20(t, J=2Hz, 1H, Ar-H);4.37(s,2H, ArCH₂N); 3.51–3.54(m, 4H);3.28–3.30(m, 2H); 3.17–3.22(m,2H);2.74(t, J=14Hz, 2H); 2.58–2.61(m, 2H);1.22–1.25(m, 3H,CH₃);1.11–1.15(m, 3H, CH₃) 3

4-[1-(3-Amino-phenyl)-1-(1-pyridin-2-ylmethyl-piperidin-4-ylidene)-methyl]-N,N-diethyl-benzamide.8.67(d, J=4.7Hz, 1H, Ar-H); 7.88(m, 1H,Ar-H); 7.51(d, J=7.4Hz, 1H,Ar-H); 7.44(m, 2H, Ar-H); 7.35(d, J=8.3Hz, 2H,Ar-H); 7.27(d, J=8.3Hz,2H, Ar-H);7.19(t, J=7.4Hz, 2H, Ar-H);7.09(s, 1H, Ar-H);4.52 (s, 2H,NCH₂Ar); 3.52(m, 2H, CH₂);3.44(m, 4H, CH₂); 3.27(m, 2H,CH₂);2.70(t,J=5.6Hz, 4H, NCH₂Me);1.12(br s, 3H, CH₃);1.10(br s, 3H, CH₃)4

4-[1-(3-Amino-phenyl)-1-(1-pyridin-4-ylmethyl-piperidin-4-ylidene)-methyl]-N,N-diethyl-benzamide.8.81(d, J=5.5Hz, 2H, Ar-H); 8.74(d,J=6.5Hz, 1H, Ar-H); 8.20(m, 2H,Ar-H);8.05(d, J=6.5Hz, 1H, Ar-H); 7.42(t,J=7.4Hz, 1H, Ar-H); 7.25(t,J=8.4Hz, 2H,Ar-H); 7.19(m, 2H, Ar-H); 6.98(m, 1H, Ar-H); 4.60(s, 2H,NCH₂Ar); 3.37(br s, 6H,CH₂); 3.18(br s, 2H, CH₂); 2.62(m, 4H,NCH₂Me);1.11(br s, 3H, CH₃); 1.00(br s,3H, CH₃) 5

4-[1-(3-Amino-phenyl)-1-(1-furan-2-ylmethyl-piperidin-4-ylidene)-methyl]-N,N-diethyl-benzamide.7.66(s, 1H, Ar-H); 7.53(t, J=7.4Hz, 1H, Ar-H); 7.35(m, 3H, Ar-H);7.27(m, 3H, Ar-H);7.22(s, 1H, Ar-H); 6.74(d, J=2.7 Hz, 1H,Ar-H); 6.52(t,J=1.8Hz, 1H, Ar-H); 4.45(s,2H, NCH₂Ar); 3.52(br s, 4H, CH₂);3.29(m, 2H,CH₂); 3.15(m, 2H, CH₂);2.75(m, 2H,NCH₂Me); 2.57(m, 2H, NCH₂Me);1.22(brs, 3H, CH₃); 1.11(br s, 3H, CH₃) 6

4-[1-(3-Amino-phenyl)-1-(1-furan-3-ylmethyl-piperidin-4-ylidene)-methyl]-N,N-diethyl-benzamide.7.70(s, 1H, Ar-H); 7.55(t, J=1.8Hz, 1H, Ar-H); 7.37(t, J=7.4Hz, 1H,Ar-H); 7.28(d,J=8.4Hz, 2H, Ar-H); 7.18(d, J=8.4Hz, 2H,Ar-H); 7.10(m, 2H,Ar-H); 6.97(s, 1H, Ar-H); 6.55(s, 1H, Ar-H); 4.17(s, 2H,NCH₂Ar); 3.48(m,4H, CH₂); 3.21(m, 2H,CH₂); 3.01(m, 2H, CH₂); 2.71(m, 2H,NCH₂Me); 2.47(m,2H, NCH₂Me);1.15(br s, 3H, CH₃); 1.03(br s, 3H, CH₃) 7

4-[1-(3-Amino-phenyl)-1-(1-thiophen-2-ylmethyl-piperidin-4-ylidene)-methyl]-N,N-diethyl-benzamide7.35(m, 2H, Ar-H); 7.22(d, J=7.5Hz, 2H,Ar-H); 7.12(d, J=8.4Hz, 2H,Ar-H); 7.07(m, 1H, Ar-H); 6.94(t, J=7.5Hz, 1H, Ar-H);6.50(m, 1H, Ar-H);6.40(br s, 1H, Ar-H);6.35(d, J=7.4Hz, 1H, Ar-H); 3.83(s, 2H,NCH₂Ar);3.44(m, 2H, CH₂); 3.31(m, 2H,CH₂); 2.74(br s, 4H, CH₂); 2.40(m,4H,NCH₂Me); 1.14(br s, 3H, CH₃); 1.02(br s,3H, CH₃) 8

4-[1-(3-Amino-phenyl)-1-(1-thiophen-3-ylmethyl-piperidin-4-ylidene)-methyl]-N,N-diethyl-benzamide7.62(d, J=5.5Hz, 1H, Ar-H); 7.34(m, 3H,Ar-H); 7:24(d, J=7.4Hz, 2H,Ar-H); 7.19(t,J=7.4Hz, 1H, Ar-H); 7. 12(dd, J=3.7, 5.6Hz,1H, Ar-H);6.83(d, J=8.4Hz, 1H, Ar-H);6.71(m, 2H, Ar-H); 4.59(s, 2H,NCH₂Ar);3.52(m, 4H, CH₂); 3.40(m, 4H, CH₂); 2.65(br s, 4H, NCH₂Me);1.22(m, 3H, CH₃);1.10(m, 6H, CH₃) 9

4-[1-(3-Amino-phenyl)-1-(1-pyrrol-2-ylmethyl-piperidin-4-ylidene)-methyl]-N,N-diethyl-benzamide1.12(br s, 3H), 1.24(br s, 3H), 2.55–2.62(m, 2H), 2.70–2.77(m, 2H),3.05–3.14(m,2H), 3.30(br s, 2H), 3.44–3.60(m, 4H),4.35(s, 2H), 6.16(s,1H), 6.37(s, 1H),6.88(s, 1H), 7.25–7.38(m, 7H), 7.52–7.56(m, 1H),10.69(s, 1H)Pharmaceutical Compositions

The novel compounds according to the present invention may beadministered orally, sublingually, intramuscularly, subcutaneously,topically, intranasally, intraperitoneally, intrathoracially,intravenously, epidurally, intrathecally, intracerebroventricularly andby injection into the joints.

A preferred route of administration is orally, intravenously orintramuscularly.

The dosage will depend on the route of administration, the severity ofthe disease, age and weight of the patient and other factors normallyconsidered by the attending physician, when determining the individualregimen and dosage level as the most appropriate for a particularpatient.

For preparing pharmaceutical compositions from the compounds of thisinvention, inert, pharmaceutically acceptable carriers can be eithersolid or liquid. Solid form preparations include powders, tablets,dispersible granules, capsules, cachets, and suppositories.

A solid carrier can be one or more substances which may also act asdiluents, flavoring agents, solubilizers, lubricants, suspending agents,binders, or tablet disintegrating agents; it can also be anencapsulating material.

In powders, the carrier is a finely divided solid which is in a mixturewith the finely divided active component. In tablets, the activecomponent is mixed with the carrier having the necessary bindingproperties in suitable proportions and compacted in the shape and sizedesired.

For preparing suppository compositions, a low-melting wax such as amixture of fatty acid glycerides and cocoa butter is first melted andthe active ingredient is dispersed therein by, for example, stirring.The molten homogeneous mixture is then poured into convenient sizedmolds and allowed to cool and solidify.

Suitable carriers are magnesium carbonate, magnesium stearate, talc,lactose, sugar, pectin, dextrin, starch, tragacanth, methyl cellulose,sodium carboxymethyl cellulose, a low-melting wax, cocoa butter, and thelike.

Salts include, but are not limited to, pharmaceutically acceptablesalts. Examples of pharmaceutically acceptable salts within the scope ofthe present invention include: acetate, benzenesulfonate, benzoate,bicarbonate, bitartrate, bromide, calcium acetate, camsylate, carbonate,chloride, citrate, dihydrochloride, edetate, edisylate, estolate,esylate, fumarate, glucaptate, gluconate, glutamate,glycollylarsanilate, hexylresorcinate, hydrabaamine, hydrobromide,hydrochloride, hydroxynaphthoate, isethionate, lactate, lactobionate,malate, maleate, mandelate, mesylate, methylbromide, methylnitrate,methylsulfate, mucate, napsylate, nitrate, pamoate (embonate),pantothenate, phosphate/diphosphate, polygalacturonate, salicylate,stearate, subacetate, succinate, sulfate, tannate, tartrate, teoclate,triethiodide, and benzathine.

Examples of pharmaceutically unacceptable salts within the scope of thepresent invention include: hydroiodide, perchlorate, tetrafluoroborate.Pharmaceutically unacceptable salts could be of use because of theiradvantageous physical and/or chemical properties, such as crystallinity.

Preferred pharmaceutically acceptable salts are hydrochlorides, sulfatesand bitartrates. The hydrochloride and sulfate salts are particularlypreferred.

The term composition is intended to include the formulation of theactive component with encapsulating material as a carrier providing acapsule in which the active component (with or without other carriers)is surrounded by a carrier which is thus in association with it.Similarly, cachets are included.

Tablets, powders, cachets, and capsules can be used as solid dosageforms suitable for oral administration.

Liquid from compositions include solutions, suspensions, and emulsions.Sterile water or water-propylene glycol solutions of the activecompounds may be mentioned as an example of liquid preparations suitablefor parenteral administration. Liquid compositions can also beformulated in solution in aqueous polyethylene glycol solution.

Aqueous solutions for oral administration can be prepared by dissolvingthe active component in water and adding suitable colorants, flavoringagents, stabilizers, and thickening agents as desired. Aqueoussuspensions for oral use can be made by dispersing the finely dividedactive component in water together with a viscous material such asnatural synthetic gums, resins, methyl cellulose, sodium carboxymethylcellulose, and other suspending agents known to the pharmaceuticalformulation art.

Preferably the pharmaceutical compositions is in unit dosage form. Insuch form, the composition is divided into unit doses containingappropriate quantities of the active component. The unit dosage form canbe a packaged preparation, the package containing discrete quantities ofthe preparations, for example, packeted tablets, capsules, and powdersin vials or ampoules. The unit dosage form can also be a capsule,cachet, or tablet itself, or it can be the appropriate number of any ofthese packaged forms.

BIOLOGICAL EVALUATION

In Vitro Model

Cell Culture

-   A. Human 293S cells expressing cloned human μ, δ, and κ receptors    and neomycin resistance were grown in suspension at 37° C. and 5%    CO₂ in shaker flasks containing calcium-free DMEM10% FBS, 5% BCS,    0.1% Pluronic F-68, and 600 μg/ml geneticin.-   B. Mouse and rat brains were weighed and rinsed in ice-cold PBS    (containing 2.5 mM EDTA, pH 7.4). The brains were homogenized with a    polytron for 15 sec (mouse) or 30 sec (rat) in ice-cold lysis buffer    (50 mM Tris, pH 7.0, 2.5 mM EDTA, with phenylmethylsulfonyl fluoride    added just prior use to 0.5 MmM from a 0.5M stock in DMSO:ethanol).    Membrane Preparation

Cells were pelleted and resuspended in lysis buffer (50 mM Tris, pH 7.0,2.5 mM EDTA, with PMSF added just prior to use to 0.1 mM from a 0.1 Mstock in ethanol), incubated on ice for 15 min, then homogenized with apolytron for 30 sec. The suspension was spun at 1000 g (max) for 10 minat 4° C. The supernatant was saved on ice and the pellets resuspendedand spun as before. The supernatants from both spins were combined andspun at 46,000 g(max) for 30 min. The pellets were resuspended in coldTris buffer (50 mM Tris/Cl, pH 7.0) and spun again. The final pelletswere resuspended in membrane buffer (50 mM Tris, 0.32 M sucrose, pH7.0). Aliquots (1 ml) in polypropylene tubes were frozen in dryice/ethanol and stored at −70° C. until use. The protein concentrationswere determined by a modified Lowry assay with SDS.

Binding Assays

Membranes were thawed at 37° C., cooled on ice, passed 3 times through a25-gauge needle, and diluted into binding buffer (50 mM Tris, 3 mMMgCl₂, 1 mg/ml BSA (Sigma A-7888), pH 1.4, which was stored at 4° C.after filtration through a 0.22 m filter, and to which had been freshlyadded 5 μg/ml aprotinin, 10 μM bestatin, 10 μM diprotin A, no DTT).Aliquots of 100 μl were added to iced 12×75 mm polypropylene tubescontaining 100 μl of the appropriate radioligand and 100 μl of testcompound at various concentrations. Total (TB) and nonspecific (NS)binding were determined in the absence and presence of 10 μM naloxonerespectively. The tubes were vortexed and incubated at 25° C. for 60–75min, after which time the contents are rapidly vacuum-filtered andwashed with about 12 ml/tube iced wash buffer (50 mM Tris, pH 7.0, 3 mMMgCl₂) through GF/B filters (Whatman) presoaked for at least 2 h in 0.1%polyethyleneimine. The radioactivity (dpm) retained on the filters wasmeasured with a beta counter after soaking the filters for at least 12 hin minivials containing 6–7 ml scintillation fluid. If the assay is setup in 96-place deep well plates, the filtration is over 96-placePEI-soaked unifilters, which were washed with 3×1 ml wash buffer, anddried in an oven at 55° C. for 2 h. The filter plates were counted in aTopCount (Packard) after adding 50 μl MS-20 scintillation fluid/well.

Functional Assays

The agonist activity of the compounds is measured by determining thedegree to which the compounds receptor complex activates the binding ofGTP to G-proteins to which the receptors are coupled. In the GTP bindingassay, GTP[γ]³⁵S is combined with test compounds and membranes fromHEK-293S cells expressing the cloned human opioid receptors or fromhomogenised rat and mouse brain. Agonists stimulate GTP[γ]³⁵S binding inthese membranes. The EC₅₀ and E_(max) values of compounds are determinedfrom dose-response curves. Right shifts of the dose response curve bythe delta antagonist naltrindole are performed to verify that agonistactivity is mediated through delta receptors.

Procedure for Rat Brain GTP

Rat brain membranes are thawed at 37° C., passed 3 times through a25-gauge blunt-end needle and diluted in the GTPyS binding (50 mM Hepes,20 M NaOH, 100 mM NaCl, 1 mM EDTA, 5 mM MgCl₂, pH 7.4, Add fresh: 1 mMDTT, 0.1% BSA). 120 μM GDP final is added membranes dilutions. The EC50and Emax of compounds are evaluated from 10-point dose-response curvesdone in 300 μl with the appropriate amount of membrane protein (20μg/well) and 100000–130000 dpm of GTPγ³⁵S per well (0.11–0.14 nM). Thebasal and maximal stimulated binding are determined in absence andpresence of 3 μM SNC-80

Data Analysis

The specific binding (SB) was calculated as TB-NS, and the SB in thepresence of various test compounds was expressed as percentage ofcontrol SB. Values of IC₅₀ and Hill coefficient (n_(H)) for ligands indisplacing specifically bound radioligand were calculated from logitplots or curve fitting programs such as Ligand, GraphPad Prism,SigmaPlot, or ReceptorFit. Values of K_(i) were calculated from theCheng-Prussoff equation. Mean±S.E.M. values of IC₅₀, K_(i) and n_(H)were reported for ligands tested in at least three displacement curves.Biological activity of the compounds of the present invention isindicated in Table 2.

TABLE 2 Biological Data. HDELTA RAT BRAIN MOUSE BRAIN (nM) (nM) (nM) Ex.# IC₅₀ EC₅₀ % EMax EC₅₀ % EMax EC₅₀ % EMax 1–8 0.22–2.18 0.55–13.493–106 5.57–106 67–155 9.08–207.5 73–144Receptor Saturation Experiments

Radioligand K_(δ) values were determined by performing the bindingassays on cell membranes with the appropriate radioligands atconcentrations ranging from 0.2 to 5 times the estimated K_(δ) (up to 10times if amiounts of radioligand required are feasible). The specificradioligand binding was expressed as pmole/mg membrane protein. Valuesof K_(δ) and B_(max) from individual experiments were obtained fromnonlinear fits of specifically bound (3) vs. nM free (F) radioligandfrom individual according to a one-site model.

Determination Of Mechano-Allodynia Using Von Frey Testing

Testing was performed between 08:00 and 16:00 h using the methoddescribed by Chaplan et al. (1994). Rats were placed in Plexiglas cageson top of a wire mesh bottom which allowed access to the paw, and wereleft to habituate for 10–15 min. The area tested was the mid-plantarleft hind paw, avoiding the less sensitive foot pads. The paw wastouched with a series of 8 Von Frey hairs with logarithmicallyincremental stiffness (0.41, 0.69, 1.20, 2.04, 3.63, 5.50, 8.51, and15.14 grams; Stoelting, Ill., USA). The von Frey hair was applied fromunderneath the mesh floor perpendicular to the plantar surface withsufficient force to cause a slight buckling against the paw, and heldfor approximately 6–8 seconds. A positive response was noted if the pawwas sharply withdrawn. Flinching immediately upon removal of the hairwas also considered a positive response. Ambulation was considered anambiguous response, and in such cases the stimulus was repeated.

Testing Protocol

The animals were tested on postoperative day 1 for the FCA-treatedgroup. The 50% withdrawal threshold was determined using the up-downmethod of Dixon (1980). Testing was started with the 2.04 g hair, in themiddle of the series. Stimuli were always presented in a consecutiveway, whether ascending or descending. In the absence of a paw withdrawalresponse to the initially selected hair, a stronger stimulus waspresented; in the event of paw withdrawal, the next weaker stimulus waschosen. Optimal threshold calculation by this method requires 6responses in the immediate vicinity of the 50% threshold, and countingof these 6 responses began when the first change in response occurred,e.g. the threshold was first crossed. In cases where thresholds felloutside the range of stimuli, values of 15.14 (normal sensitivity) or0.41 (maximally allodynic) were respectively assigned. The resultingpattern of positive and negative responses was tabulated using theconvention, X=no withdrawal; O=withdrawal, and the 50% withdrawalthreshold was interpolated using the formula:50% g threshold=10^((Xf+kδ))/10,000where Xf=value of the last von Frey hair used (log units); k=tabularvalue (from Chaplan et al. (1994)) for the pattern of positive/negativeresponses; and δ=mean difference between stimuli (log units). Hereδ=0.224.

Von Frey thresholds were converted to percent of maximum possible effect(% MPE), according to Chaplan et al. 1994. The following equation wasused to compute % MPE:

${\%\mspace{14mu}{MPE}} = {\frac{{{Drug}\mspace{14mu}{treated}\mspace{14mu}{threshold}\mspace{14mu}(g)} - {{allodynia}\mspace{14mu}{threshold}\mspace{14mu}(g)}}{{{Control}\mspace{14mu}{threshold}\mspace{14mu}(g)} - {{allodynia}\mspace{14mu}{threshold}\mspace{14mu}(g)}} \times 100}$Administration Of Test Substance

Rats were injected (subcutaneously, intraperitoneally, intravenously ororally) with a test substance prior to von Frey testing, the timebetween administration of test compound and the von Frey test varieddepending upon the nature of the test compound.

Writhing Test

Acetic acid will bring abdominal contractions when administeredintraperitoneally in mice. These will then extend their body in atypical pattern. When analgesic drugs are administered, this describedmovement is less frequently observed and the drug selected as apotential good candidate.

A complete and typical Writhing reflex is considered only when thefollowing elements are present: the animal is not in movement; the lowerback is slightly depressed; the plantar aspect of both paws isobservable. In this assay, compounds of the present inventiondemonstrate significant inhibition of writhing responses after oraldosing of 1–100 μmol/kg.

(i) Solutions Preparation

Acetic acid (AcOH): 120 μL of Acetic Acid is added to 19.88 ml ofdistilled water in order to obtain a final volume of 20 ml with a finalconcentration of 0.6% AcOH. The solution is then mixed (vortex) andready for injection.

Compound (drug): Each compound is prepared and dissolved in the mostsuitable vehicle according to standard procedures.

(ii) Solutions Administration

The compound (drug) is administered orally, intraperitoneally (i.p.),subcutaneously (s.c.) or intravenously (i.v.)) at 10 ml/kg (consideringthe average mice body weight) 20, 30 or 40 minutes (according to theclass of compound and its characteristics) prior to testing. When thecompound is delivered centrally: Intraventricularly (i.c.v.) orintrathecally (i.t.) a volume of 5 μL is administered.

The AcOH is administered intraperitoneally (i.p.) in two sites at 10ml/kg (considering the average mice body weight) immediately prior totesting.

(iii) Testing

The animal (mouse) is observed for a period of 20 minutes and the numberof occasions (Writhing reflex) noted and compiled at the end of theexperiment. Mice are kept in individual “shoe box” cages with contactbedding. A total of 4 mice are usually observed at the same time: onecontrol and three doses of drug.

For the anxiety and anxiety-like indications, efficacy has beenestablished in the geller-seifter conflict test in the rat.

For the functional gastrointestina disorder indication, efficacy can beestablished in the assay described by Coutinho SV et al, in AmericanJournal of Physiology—Gastrointestinal & Liver Physiology.282(2):G307-16, 2002 Feb, in the rat.

1. A compound of the formula I

wherein R¹ is selected from any one of

where each R¹ phenyl ring and R¹ heteroaromatic ring may independentlybe further substituted by 1, 2 or 3 substituents independently selectedfrom straight and branched C₁–C₆ alkyl, NO₂, CF₃, C₁–C₆ alkoxy, chioro,fluoro, bromo, and jodo, as well as salts thereof.
 2. A compoundaccording to claim 1, wherein each R¹ heteroaromatic ring mayindependently be further substituted by 1,2 or 3 substituentsindependently selected from methyl, CF₃, chloro, fluoro, bromo, andiodo.
 3. A compound according to claim 1, wherein each R¹heteroaromnatic ring may independently be further substituted by amethyl group.
 4. A compound according to claim 1, wherein R¹ pyridinyl,thienyl or furanyl.
 5. A compound according to claim 1, selected fromany one of:4-[1-(3-Amino-phenyl)-1-(1-pyridin-2-ylmethyl-piperidiu-4-ylidene)-methyl]-N,N-diethyl-benzamide,4-[1-(3-Amino-phenyl)-1-(1-pyridin-4-ylmethyl-piperidin-4-ylidene)-methyl]-N,N-diethyl-benzamide,4-[1-(3-Amino-phenyl)-1-(1-furan-2-ylmethyl-piperidin-4-ylidene)-methyl]-N,N-diethyl-benzamide,4-[1-(3-Amino-phenyl)-1-(1-furan-3-ylmethyl-piperidin-4-ylidene)-methyl]-N,N-diethyl-benzamide;4-[1-(3-Amino-phenyl)-1-(1-thiophen-2-ylmethyl-piperidin-4-ylidene)-methyl]-N,N-diethyl-benzamide,4-[1-(3-Amino-phenyl)-1-(1-pyrrol-2-ylmethyl-piperidin-4-ylidene)-methyl)]-N,N-diethyl-benzamide,and4-[1-(3-Amino-phenyl)-1-(1-thiophen-3-ylmethyl-piperidin-4-ylidene)-methyl]-N,N-diethyl-benzamide.6. A compound according to any of the preceding claims, in form of itshydrochloride, dihydrochioride, sulfate, tartratc, ditrifluoroacetate orcitrate salts.
 7. A process for preparing a compound of formula I,

comprising the reacting a compound of the general formula II

wherein PG is selected from Boc, CBZ, a benzyl and 2,4-diniethoxybenzyl,with 3-aminophenyl boronic acid, using a palladium catalyst in thepresence of a base to give the compounds of general formula III,

which is thereafter deprotected, under standard conditions and alkylatedunder reductive conditions with a compound of the general formula R¹-CHOto give compounds of the general formula I, wherein R¹ is selected fromany one of pyridinyl, thienyl, furanyl; imidazolyl, triazolyl, pyrrolyl,thiazolyl and pyridyl-N-oxide, wherein each R¹ heteroarornatic ring mayindependently be further substituted by 1, 2 or 3 substituentsindependently selected from straight and branched C₁–C₆ alkyl, NO₂, CF₃,C₁–C₆ alkoxy, chloro, fluoro, bromo, and iodo, as well as salts thereof.8. A pharmaceutical composition comprising a compound of the formula Iaccording to claim 1 as an active ingredient, together with apharmaceutically acceptable carrier.
 9. A method for the treatment ofpain, whereby an effective amount of a compound of the formula Iaccording to claim 1 is administered to a subject in need of painmanagement.
 10. A method for the treatment of anxiety, whereby aneffective amount of a compound of the formula I according to claim 1, isadministered to a subject suffering from said anxiety.